PROJECT SUMMARY Based on the well-documented relationship that exists between ovarian function and women?s health, for decades hormone therapy was considered by many as the standard-of-care for the management, and even the prevention, of post-menopausal health complications. However, recent clinical trials have raised concerns over hormone therapy and increased risks for coronary heart disease, stroke, blood clots, and cancer. While the general thinking behind hormone therapy ? viz. provision of ovarian hormones lost at menopause would serve as a ?replacement? for failed ovarian function ? seems reasonable, it is perhaps not surprising that replacement of only one (estrogen) or two (estrogen-progestin) of the multitude of bioactive factors produced by the ovaries during reproductive life would ultimately prove insufficient. We have been working on the idea that ?ovarian therapy? would serve as a far better ?replacement? for alleviating health consequences of failed ovarian function with age. We eventually generated a mouse model that maintains an adequate reserve of oocyte-containing follicles, and an ensuing continuation of ovarian function, well into advanced chronological ages. Long-term follow-up work showed this strategy indeed yielded immense health benefits in aging females without an increase in cancer in any tissue. While we were excited to demonstrate this key proof-of-concept in mammals, there was a major downside: the ?gene knockout? approach used was not amenable for translation to women. Nonetheless, the work underscored the importance of continuing efforts to elucidate mechanisms underlying endowment and depletion of oocyte-containing follicles in the context of ovarian aging. This led us to discover the existence of female germline or oogonial stem cells (OSCs) in mice and then in humans, and the role these cells play in supporting ovarian function during adulthood. Of the many new areas this paradigm shift opened, one of the most exciting revolves around the use of regenerative medicine to extend functional ovarian lifespan into later ages of life. Achieving this, however, will require a detailed understanding of cues that drive OSC differentiation, and how aging impacts on these cues such that the ability of ovaries to sustain their oocyte reserves becomes compromised with age. To this end we found that application of mechanical ?strain? to OSCs activates their differentiation into oocytes. We also found that ovarian matrix proteins, which directly influence biomechanical properties of tissue, not only decline with age but also serve as direct activators of OSC differentiation. Here we have designed a number of in-vitro and in-vivo studies to rigorously test the hypothesis that progressive loss of mouse and human OSC function in ovaries with age is directly tied to a reduction in biomechanical stimulatory signaling occurring concomitant with aging-associated changes in matrix proteins. Completion of these studies will both support our central hypothesis as well as more broadly highlight the significance of mechanotransduction to adult stem cell function in the context of organ failure with age.